PROJECT SUMMARY Cardiovascular disease is common and deadly. While lower organisms have a tremendous capacity for regeneration, the adult mammalian heart is more limited in its capacity for regeneration and remuscularization of the damaged tissue following an insult such as a myocardial infarction. Recently, we defined an evolutionary conserved Shh-Gli1-Mycn cascade in newt, mouse and human that promotes cardiomyocyte proliferation in vitro and cardiac repair. These studies used pharmacological agents, genetic mouse models, viral vectors, fate mapping techniques, molecular biological technologies, and time lapse videomicroscopy. Importantly, the results of these studies and these tools and reagents provide a platform for the proposed studies that will dissect the mechanisms that govern cardiovascular proliferation. Therefore, our overall hypothesis is that the Shh downstream effector, Mycn, regulates cardiomyocyte and endothelial cellular proliferation. In these proposed studies, we will use a number of novel genetic models that we have engineered, bioinformatics algorithms that we developed and we take an innovative approach to dissect the role of Mycn as an important factor that governs cardiovascular proliferation. To examine our hypotheses, we will address the following specific aims: Specific Aim #1: To define the role of Mycn in the endothelial and cardiomyocyte lineages; Specific Aim #2: To define the impact of Mycn overexpression in cardiovascular lineages and Specific Aim #3: To define the Mycn targets in the cardiomyocyte lineage. These aims will utilize our recently engineered genetic mouse models, inducible viral vectors, modified mRNA- GFP constructs, ATAC-seq, MNase-seq, ChIP-seq, and bioinformatics algorithms to comprehensively define the role for Mycn as an essential factor that governs cardiomyocyte and endothelial cell proliferation and will promote heart regeneration following injury. Given the tremendous morbidity and mortality of cardiovascular disease in our society, the potential impact of this proposal is tremendous.